棉花幼苗对低温胁迫的响应及抗冷机制初步研究
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摘要
【目的】探讨棉花不同组织对低温胁迫的响应,初步探讨研究抗冷的生理生化和分子机制。【方法】测定了4个抗冷性差异显著的材料在低温处理后的生物量、抗氧化酶活力和可溶性蛋白含量,同时分析了抗冷相关基因在豫2067根、茎、叶的表达情况。【结果】4℃低温处理24 h对冷敏感材料生长抑制最大,对根系生长的抑制大于地上部分;细胞膜透性测定结果表明,受低温胁迫后4个材料根系细胞膜能保持相对稳定的结构,而叶片细胞膜对低温胁迫敏感,抗冷材料叶片细胞膜稳定性大于冷敏感材料,低温胁迫后叶片细胞膜透性与棉花的抗冷性呈负相关,可以作为棉花抗冷性的鉴定指标;棉花在受到低温胁迫24 h后,抗冷材料根中SOD的活力基本不变,冷敏感材料根中SOD的活力却极显著下降,2个材料的根系中POD、CAT的活力均下降,但抗冷材料豫2067下降的幅度小于冷敏感材料,可溶性蛋白含量在抗冷材料叶茎中基本不变,在冷敏材料的叶片和茎中均下降。低温胁迫后5个与抗冷相关的基因在豫2067叶片中上调表达的多于下调表达的,上调表达的基因分别是脂肪酸脱氢酶基因、胁迫诱导蛋白基因、假定R2R3-MYB转录因子基因、b HLH1转录因子基因;在根中下调表达的多于上调表达的,下调的基因分别为b HLH1转录因子基因、胁迫诱导蛋白基因、伸展蛋白基因2,这与根系的生长受抑制的结果是一致的。【结论】推测这些抗冷相关基因在叶片中的上调表达与叶片生长受低温抑制程度低有一定的关系。
[Objective] Our research aimed to investigate the effects of low temperature on tissues of cotton and the physiological,biochemical and molecular mechanisms of response to low temperature.[Method] We measured biomass,antioxidant enzyme activity and soluble protein content of four cotton materials with significantly different chilling tolerances after 4 ℃ low temperature treatment.We also measured the relative expression of chilling resistance-related genes of different tissues of cotton cultivar Yu 2067 after 4 ℃ treatment.[Result] The low temperature treatment(4 ℃ for 24 h) resulted in greater growth inhibition of chilling-sensitive than chilling-tolerant materials,with inhibition of root growth higher than that of aerial parts.According to cell membrane permeability measurements,root cell membranes of the four tested cottons were relatively stable under low temperature stress.Compared with membranes of roots and stems,leaf cell membranes were more sensitive to low temperature stress.Leaf cell membranes of chilling-resistant cotton materials were more stable than those of chilling-sensitive ones.After low temperature stress,the cell membrane permeability of cotton leaves was negatively correlated with cold resistance,suggesting the use of this parameter as an indicator of cold resistance in cotton.SOD activity in the roots of chilling-resistant materials remained unchanged,whereas that of cold sensitive-ones decreased extremely significantly.POD and CAT activities decreased in the roots of two cotton materials after low temperature stress.The activities of two types of antioxidant enzymes declined less markedly in cold-resistant Yu 2067 than in cold-sensitive materials.The soluble protein content of leaves and stems of cold-resistant cotton remained basically unchanged but decreased in cold-sensitive materials.Expression analysis of five chilling tolerance-related genes in Yu 2067 revealed that more genes were up-regulated than down-regulated in leaves.The four genes up-regulated in leaves were 3FAD(fatty acid desaturase),stress-induced protein,putative R2R3-MYB transcription factor and b HLH1 transcription factor.In contrast,more genes were down-regulated than up-regulated in roots.In roots,the three down-regulated genes were b HLH1 transcription factor,stress-induced protein and expansin 2.These expression patterns were consistent with the observed root growth inhibition.[Conclusion] Our results imply that a relationship exists between up-regulation of the four cold resistance-related genes in leaves and inhibition of leaf growth under low temperature conditions.
[Objective] Our research aimed to investigate the effects of low temperature on tissues of cotton and the physiological,biochemical and molecular mechanisms of response to low temperature.[Method] We measured biomass,antioxidant enzyme activity and soluble protein content of four cotton materials with significantly different chilling tolerances after 4 ℃ low temperature treatment.We also measured the relative expression of chilling resistance-related genes of different tissues of cotton cultivar Yu 2067 after 4 ℃ treatment.[Result] The low temperature treatment(4 ℃ for 24 h) resulted in greater growth inhibition of chilling-sensitive than chilling-tolerant materials,with inhibition of root growth higher than that of aerial parts.According to cell membrane permeability measurements,root cell membranes of the four tested cottons were relatively stable under low temperature stress.Compared with membranes of roots and stems,leaf cell membranes were more sensitive to low temperature stress.Leaf cell membranes of chilling-resistant cotton materials were more stable than those of chilling-sensitive ones.After low temperature stress,the cell membrane permeability of cotton leaves was negatively correlated with cold resistance,suggesting the use of this parameter as an indicator of cold resistance in cotton.SOD activity in the roots of chilling-resistant materials remained unchanged,whereas that of cold sensitive-ones decreased extremely significantly.POD and CAT activities decreased in the roots of two cotton materials after low temperature stress.The activities of two types of antioxidant enzymes declined less markedly in cold-resistant Yu 2067 than in cold-sensitive materials.The soluble protein content of leaves and stems of cold-resistant cotton remained basically unchanged but decreased in cold-sensitive materials.Expression analysis of five chilling tolerance-related genes in Yu 2067 revealed that more genes were up-regulated than down-regulated in leaves.The four genes up-regulated in leaves were 3FAD(fatty acid desaturase),stress-induced protein,putative R2R3-MYB transcription factor and b HLH1 transcription factor.In contrast,more genes were down-regulated than up-regulated in roots.In roots,the three down-regulated genes were b HLH1 transcription factor,stress-induced protein and expansin 2.These expression patterns were consistent with the observed root growth inhibition.[Conclusion] Our results imply that a relationship exists between up-regulation of the four cold resistance-related genes in leaves and inhibition of leaf growth under low temperature conditions.
引文
[1]Godfray H C J.Food security:the challenge of feeding 9 billion people[J].Science,2010,327(5967):812-818.
[2]David T,Christian B,Jason H,et al.Global food demand and the sustainable intensification of agriculture[J].Proceedings of the National Academy of Sciences of the United States of America,2011,108(50):20260-20264.
[3]李新国,毕玉平,赵世杰,等.短时低温胁迫对甜椒叶绿体超微结构和光系统的影响[J].中国农业科学,2005,38(6):1226-1231.Li Xinguo,Bi Yuping,Zhao Shijie,et al.Effects of short-term chilling stress on the photosystems and chloroplast ultrastructure in sweet pepper[J].Scientia Agricultura Sinica,2005,38(6):1226-1231.
[4]利容千,王建波.植物逆境细胞及生理学[M].武汉:武汉大学出版社,2002:141.Li Rongqian,Wang Jianbo.Plant stress cell and physiology[M].Wuhan:Wuhan University Press,2002:141.
[5]Li Xinguo,Meng Qinwei,Jiang Guoqiang,et al.The susceptibility of cucumber and sweet pepper to chilling under low irradiance is related to energy dissipation and water-water cycle[J].Photosynthetica,2003,41(2):259-265.
[6]Barrero-Gil J,Salinas J.Post-translational regulation of cold acclimation response[J].Plant Science,2013,205/206:48-54.
[7]Guy C,Haskell D,Li Qinbao.Association of proteins with the stress 70 molecular chaperones at low temperature:evidence for the existence of cold labile proteins in spinach[J].Cryobiology,1998,36(4):301-314.
[8]Haldimann P.Effects of changes in growth temperature on photosynthesis and carotenoid composition in Zea mays leaves[J].Physiologia Plantarum,1996,97:554-562.
[9]潘瑞炽,董愚得.植物生理学[M].北京:高等教育出版社,1995:322-328.Pan Ruichi,Dong Yude.Plant physiology[M].Beijing:Higher Education Press,1995:322-328.
[10]董合林,王润珍,李鹏程,等.不同施氮水平及氮磷钾肥配施对棉花产量与氮肥利用率的影响[C]//中国棉花学会2010年年会论文汇编.安阳:中国棉花杂志社,2010:285.Dong Helin,Wang Runzhen,Li Pengcheng,et al.Effect of N application rates and N,P and K combinations on cotton yield and utilization[C]//China Society of Cotton Sci-Tech Proceedings of 2010 Annual Meeting of CSCS.Anyang:China Cotton Magazine House,2010:285.
[11]王俊娟,王德龙,樊伟莉,等.陆地棉萌发至三叶期不同生育阶段耐盐特性研究[J].生态学报,2011,31(13):3720-3727.Wang Junjuan Wang Delong,Fan Weili,et al.Study on the characters of salt-tolerance at different growth stages in cotton[J].Acta Ecologica Sinica,2011,31(13):3720-3727.
[12]李合生.植物生理生化实验原理和技术[M].北京:高等教育出版社,2000:148-219.Li Hesheng.Plant physiology and biochemistry experimental principles and techniques[M].Beijing:Higher Education Press,2000:148-219.
[13]Afrin S,Zhu Jie,Cao Hongzhe,et al.Molecular cloning and expression profile of an abiotic stress and hormone responsive MYB transcription factor gene from Panax ginseng[J].Acta Biochimica et Biophysica Sinica,2015,47(4):267-277.
[14]Wan Xianchong,Zwiazek J J,Lieffers V J,et al.Hydraulic conductance in aspen(Populus tremuloides)seedlings exposed to low root temperatures[J].Tree Physiology,2001,21(10):691-696.
[15]Tyree M T,Nardini A,Salleo S,et al.The dependence of leaf hydraulic conductance on irradiance during HPFM measurements:any role for stomatal response?[J].Journal of Experimental Botany,2005,56(412):737-744.
[16]Lee S H,Chung G C,Jang J Y,et al.Overexpression of PIP2;5aquaporin alleviates effects of low root temperature on cell hydraulic conductivity and growth in Arabidopsis[J].Plant Physiology,2012,159(1):479-488.
[17]Bauerfeind M A,Winkelmann T,Franken P,et al.Transcriptome,carbohydrate,and phytohormone analysis of Petunia hybrida reveals a complex disturbance of plant functional integrity under mild chilling stress[J/OL].Frontiers in Plant Science,2015,6:583[2016-03-09].http://journal.frontiersin.org/article/10.3389/fpls.2015.00583/pdf.DOI:10.3389/fpls.2015.00583.
[18]Liu Yajie,Jiang Haifeng,Zhao Zhiguang,et al.Nitric oxide synthase like activity-dependent nitric oxide production protects against chilling-induced oxidative damage in Chorispora bungeana suspension cultured cells[J].Plant Physiology and Biochemistry,2010,48(12):936-944.
[19]田景花,王红霞,张志华,等.低温逆境对不同核桃品种抗氧化系统及超微结构的影响[J].应用生态学报,2015,26(5):1320-1326.Tian Jinghua,Wang Hongxia,Zhang Zhihua,et al.Effects of chilling stress on antioxidant system and ultrastructure of walnut cultivars[J].Chinese Journal of Applied Ecology,2015,26(5):1320-1326.
[20]Caffagni A,Pecchioni N,Francia E,et al.Candidate gene expression profiling in two contrasting tomato cultivars under chilling stress[J].Biologia Plantarum,2014,58(2):283-295.
[21]Zhou Mingqi,Xu Ming,Wu Lihua,et al.Cb CBF from Capsella bursa-pastoris enhances cold tolerance and restrains growth in Nicotiana tabacum by antagonizing with gibberellin and affecting cell cycle signaling[J].Plant Molecular Biology,2014,85(3):259-275.
[22]Steponkus P L.Role of plasma membrane in freezing injury and cold acclimation[J].Annual Review of Plant Biology,1984,35:543-584.
[23]Upchurch R G.Fatty acid unsaturation,mobilization,and regulation the response of plants to stress[J].Biotechnology Letters,2008,30:967-977.
[24]He Jing,Yang Zhaojie,Hu Binbin,et al.Correlation of polyunsaturated fatty acids with the cold adaptation of Rhodotorula glutinis[J].Yeast,2015,32(11):683-690.
[25]Shi Jinlei,Cao Yingping,Fan Xiaorong,et al.A rice microsomal delta-12 fatty acid desaturase can enhance resistance to cold stress in yeast and Oryza sativa[J].Molecular Breeding,2012,29(3):743-757.
[26]周洲.转脂肪酸去饱和酶基因Pt FAD2和Pt FAD3银腺杨84K的抗寒性研究[D].北京:中国林业科学研究院,2007.Zhou Zhou.The cold tolerance of transgenic Populus alba×Populus glandulossa 84K with fatty acid desaturase genes PtFAD2 and Pt FAD3[D].Beijing:Chinese Academy of Forestry,2007.
[27]Cho H T,Cosgrove D J.Regulation of root hair initiation and expansin gene expression in Arabidopsis[J].Plant Cell,2002,14(12):3237-3253.
[28]Wu Y J,Thorne E T,Sharp R E.Modification of expansin transcript levels in the maize primary root at low water potentials[J].Plant Physiology,2001,126(4):1471-1479.
[2]David T,Christian B,Jason H,et al.Global food demand and the sustainable intensification of agriculture[J].Proceedings of the National Academy of Sciences of the United States of America,2011,108(50):20260-20264.
[3]李新国,毕玉平,赵世杰,等.短时低温胁迫对甜椒叶绿体超微结构和光系统的影响[J].中国农业科学,2005,38(6):1226-1231.Li Xinguo,Bi Yuping,Zhao Shijie,et al.Effects of short-term chilling stress on the photosystems and chloroplast ultrastructure in sweet pepper[J].Scientia Agricultura Sinica,2005,38(6):1226-1231.
[4]利容千,王建波.植物逆境细胞及生理学[M].武汉:武汉大学出版社,2002:141.Li Rongqian,Wang Jianbo.Plant stress cell and physiology[M].Wuhan:Wuhan University Press,2002:141.
[5]Li Xinguo,Meng Qinwei,Jiang Guoqiang,et al.The susceptibility of cucumber and sweet pepper to chilling under low irradiance is related to energy dissipation and water-water cycle[J].Photosynthetica,2003,41(2):259-265.
[6]Barrero-Gil J,Salinas J.Post-translational regulation of cold acclimation response[J].Plant Science,2013,205/206:48-54.
[7]Guy C,Haskell D,Li Qinbao.Association of proteins with the stress 70 molecular chaperones at low temperature:evidence for the existence of cold labile proteins in spinach[J].Cryobiology,1998,36(4):301-314.
[8]Haldimann P.Effects of changes in growth temperature on photosynthesis and carotenoid composition in Zea mays leaves[J].Physiologia Plantarum,1996,97:554-562.
[9]潘瑞炽,董愚得.植物生理学[M].北京:高等教育出版社,1995:322-328.Pan Ruichi,Dong Yude.Plant physiology[M].Beijing:Higher Education Press,1995:322-328.
[10]董合林,王润珍,李鹏程,等.不同施氮水平及氮磷钾肥配施对棉花产量与氮肥利用率的影响[C]//中国棉花学会2010年年会论文汇编.安阳:中国棉花杂志社,2010:285.Dong Helin,Wang Runzhen,Li Pengcheng,et al.Effect of N application rates and N,P and K combinations on cotton yield and utilization[C]//China Society of Cotton Sci-Tech Proceedings of 2010 Annual Meeting of CSCS.Anyang:China Cotton Magazine House,2010:285.
[11]王俊娟,王德龙,樊伟莉,等.陆地棉萌发至三叶期不同生育阶段耐盐特性研究[J].生态学报,2011,31(13):3720-3727.Wang Junjuan Wang Delong,Fan Weili,et al.Study on the characters of salt-tolerance at different growth stages in cotton[J].Acta Ecologica Sinica,2011,31(13):3720-3727.
[12]李合生.植物生理生化实验原理和技术[M].北京:高等教育出版社,2000:148-219.Li Hesheng.Plant physiology and biochemistry experimental principles and techniques[M].Beijing:Higher Education Press,2000:148-219.
[13]Afrin S,Zhu Jie,Cao Hongzhe,et al.Molecular cloning and expression profile of an abiotic stress and hormone responsive MYB transcription factor gene from Panax ginseng[J].Acta Biochimica et Biophysica Sinica,2015,47(4):267-277.
[14]Wan Xianchong,Zwiazek J J,Lieffers V J,et al.Hydraulic conductance in aspen(Populus tremuloides)seedlings exposed to low root temperatures[J].Tree Physiology,2001,21(10):691-696.
[15]Tyree M T,Nardini A,Salleo S,et al.The dependence of leaf hydraulic conductance on irradiance during HPFM measurements:any role for stomatal response?[J].Journal of Experimental Botany,2005,56(412):737-744.
[16]Lee S H,Chung G C,Jang J Y,et al.Overexpression of PIP2;5aquaporin alleviates effects of low root temperature on cell hydraulic conductivity and growth in Arabidopsis[J].Plant Physiology,2012,159(1):479-488.
[17]Bauerfeind M A,Winkelmann T,Franken P,et al.Transcriptome,carbohydrate,and phytohormone analysis of Petunia hybrida reveals a complex disturbance of plant functional integrity under mild chilling stress[J/OL].Frontiers in Plant Science,2015,6:583[2016-03-09].http://journal.frontiersin.org/article/10.3389/fpls.2015.00583/pdf.DOI:10.3389/fpls.2015.00583.
[18]Liu Yajie,Jiang Haifeng,Zhao Zhiguang,et al.Nitric oxide synthase like activity-dependent nitric oxide production protects against chilling-induced oxidative damage in Chorispora bungeana suspension cultured cells[J].Plant Physiology and Biochemistry,2010,48(12):936-944.
[19]田景花,王红霞,张志华,等.低温逆境对不同核桃品种抗氧化系统及超微结构的影响[J].应用生态学报,2015,26(5):1320-1326.Tian Jinghua,Wang Hongxia,Zhang Zhihua,et al.Effects of chilling stress on antioxidant system and ultrastructure of walnut cultivars[J].Chinese Journal of Applied Ecology,2015,26(5):1320-1326.
[20]Caffagni A,Pecchioni N,Francia E,et al.Candidate gene expression profiling in two contrasting tomato cultivars under chilling stress[J].Biologia Plantarum,2014,58(2):283-295.
[21]Zhou Mingqi,Xu Ming,Wu Lihua,et al.Cb CBF from Capsella bursa-pastoris enhances cold tolerance and restrains growth in Nicotiana tabacum by antagonizing with gibberellin and affecting cell cycle signaling[J].Plant Molecular Biology,2014,85(3):259-275.
[22]Steponkus P L.Role of plasma membrane in freezing injury and cold acclimation[J].Annual Review of Plant Biology,1984,35:543-584.
[23]Upchurch R G.Fatty acid unsaturation,mobilization,and regulation the response of plants to stress[J].Biotechnology Letters,2008,30:967-977.
[24]He Jing,Yang Zhaojie,Hu Binbin,et al.Correlation of polyunsaturated fatty acids with the cold adaptation of Rhodotorula glutinis[J].Yeast,2015,32(11):683-690.
[25]Shi Jinlei,Cao Yingping,Fan Xiaorong,et al.A rice microsomal delta-12 fatty acid desaturase can enhance resistance to cold stress in yeast and Oryza sativa[J].Molecular Breeding,2012,29(3):743-757.
[26]周洲.转脂肪酸去饱和酶基因Pt FAD2和Pt FAD3银腺杨84K的抗寒性研究[D].北京:中国林业科学研究院,2007.Zhou Zhou.The cold tolerance of transgenic Populus alba×Populus glandulossa 84K with fatty acid desaturase genes PtFAD2 and Pt FAD3[D].Beijing:Chinese Academy of Forestry,2007.
[27]Cho H T,Cosgrove D J.Regulation of root hair initiation and expansin gene expression in Arabidopsis[J].Plant Cell,2002,14(12):3237-3253.
[28]Wu Y J,Thorne E T,Sharp R E.Modification of expansin transcript levels in the maize primary root at low water potentials[J].Plant Physiology,2001,126(4):1471-1479.